Recently, nonisotopic (often automated) immunoassays for measuring serum free thyroxin (FT4) have become available. Though more costly then radioimmunoassays, they are considerably more convenient. We studied the influence of endogenous albumin and thyroxin-binding globulin concentration on five automated, nonisotopic methods of measuring FT4 (Enzymun on ES300 [one-step], Stratus I and II [one-and-a-half-step], Delfia [two-step], and IMx [two-step]). We observed that they (a) are influenced very little by endogenous serum binding proteins and (b) seem to have sufficient within-run precision to justify performing single measurements on patient specimens. Additional studies are now in progress to assess the diagnostic performance of FT4 methods based on the use of radiolabeled T4 or T4 analogs in patients with non-thyroidal illnesses.